title: Novel mechanism of hexamer ring assembly in protein/RNA interactions revealed by single molecule imaging.
abstract: Many nucleic acid-binding proteins and the AAA+ family form hexameric rings, but the mechanism of hexamer assembly is unclear. It is generally believed that the specificity in protein/RNA interaction relies on molecular contact through a surface charge or 3D structure matching via conformational capture or induced fit. The pRNA of bacteriophage phi29 DNA-packaging motor also forms a ring, but whether the pRNA ring is a hexamer or a pentamer is under debate. Here, single molecule studies elucidated a mechanism suggesting the specificity and affinity in protein/RNA interaction relies on pRNA static ring formation. A combined pRNA ring-forming group was very specific for motor binding, but the isolated individual members of the ring-forming group bind to the motor nonspecifically. pRNA did not form a ring prior to motor binding. Only those RNAs that formed a static ring, via the interlocking loops, stayed on the motor. Single interlocking loop interruption resulted in pRNA detachment. Extension or reduction of the ring circumference failed in motor binding. This new mechanism was tested by redesigning two artificial RNAs that formed hexamer and packaged DNA. The results confirmed the stoichiometry of pRNA on the motor was the common multiple of two and three, thus, a hexamer.
authors.affiliation: Department of Biomedical Engineering, College of Engineering/College of Medicine, University of Cincinnati, Cincinnati, OH 45221, USA.
authors.forename: Feng
authors.initials: F
authors.lastname: Xiao
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.forename: Peixuan
authors.initials: P
authors.lastname: Guo
mesh_terms: D017103:Bacillus Phages / Q000235:genetics* / Q000648:ultrastructure; D001483:Base Sequence; D002213:Capsid / Q000737:chemistry / Q000648:ultrastructure; D036022:Capsid Proteins / Q000378:metabolism*; D042003:DNA Packaging; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D012367:RNA, Viral / Q000737:chemistry* / Q000648:ultrastructure; D016601:RNA-Binding Proteins / Q000378:metabolism; D009626:Terminology as Topic; D014771:Virion / Q000502:physiology; D019065:Virus Assembly
chemical_list: D036022:Capsid Proteins; D012367:RNA, Viral; D016601:RNA-Binding Proteins; C097975:portal protein, bacteriophage phi29